Journal: Frontiers in Immunology

Article Title: Ligand-based targeting of c-kit using engineered γδ T cells as a strategy for treating acute myeloid leukemia

doi: 10.3389/fimmu.2023.1294555

Figure Lengend Snippet: Treatment of hSCF sBite-modified γδ T cells only moderately prolongs survival in vivo , despite aggressive treatment regimen. (A) Experimental design. Briefly, NSG mice were pre-conditioned with 20 mg/kg busulfan IP on day -1, then injected with 5 × 10 6 CMK cells via tail-vein injection in the morning on day 0. Beginning in the afternoon on day 0, and then once daily for the next 3 days for a total of 4 doses, 1 × 10 7 γδ T cells were injected via tail-vein injection. Mice were subjected to bioluminescence imaging for the following 3 weeks, then followed for survival until they met endpoint. n = 8 untreated, n = 6 mock T treated, n = 6 hSCF sBite treated, n = 3 mSCF CAR treated. (B) Bioluminescence images. (C) Peripheral blood leukocytes were collected 3 weeks after the start of treatment and assessed for presence of CD33 + CMK cells. Error bars represent SD. Statistical analysis represents Student’s t test (ns, p > 0.05). (D) MFI of c-kit on CMK cells within the periphery. Statistical analysis represents Student’s t test (ns, p > 0.05). (E) Kaplan-Meier survival analysis. Untreated and mock γδ T treated groups were combined as a control group. Statistical analysis represents log rank (Mantel-Cox) test (ns > 0.05). P-value is shown. (F) Representative flow plots of hCD33 + hCD45 + CMK cells in the bone marrow of an untreated mouse sacrificed near end-point.

Article Snippet: Bioluminescence was quantified using Living Image Software (PerkinElmer, Waltham, MA, USA) or Aura Software (Spectral Instruments Imaging, Tucson, AZ, USA).

Techniques: Modification, In Vivo, Injection, Imaging, Control

Journal: iScience

Article Title: Defining cis-regulatory elements and transcription factors that control human cortical interneuron development

doi: 10.1016/j.isci.2024.109967

Figure Lengend Snippet:

Article Snippet: Chemidoc Image Lab Touch Software , Bio-Rad , https://www.bio-rad.com/en-us/product/chemidoc-mp-imaging-system?ID=NINJ8ZE8Z.

Techniques: Recombinant, Luciferase, Real-time Polymerase Chain Reaction, Plasmid Preparation, Software

Journal: Cells

Article Title: ‘Shared-Hook’ and ‘Changed-Hook’ Binding Activities of Herpesviral Core Nuclear Egress Complexes Identified by Random Mutagenesis

doi: 10.3390/cells11244030

Figure Lengend Snippet: Qualitative and quantitative analyses of HCMV AD169-GFP ΔUL53 virus reconstitution on the different recombinant HFF cell populations. HFFs that express pUL53, pUL53-Flag or pUL53::sHook1-Flag were transfected with the infectious bacterial artificial chromosome of HCMV ΔUL53. ( A ) Images of the GFP signals and the respective brightfield illuminations were taken at indicated time points; scale bar in panel A, picture 1 marks 500 μm. ( B ) Quantitative analysis of fluorescence-positive cells was achieved with automated counting by the CellReporterXpress ® software (Molecular Devices LLC, San Jose, CA, USA). Counting parameters for positive nuclei were set to an intensity of at least 100, a minimal width of 7 and a maximal width of 30. Measurements were performed in biological sextuplicates per cell population of 5.05% area of the well and mean values ± SD are given. ( C ) qPCR-based assay for the determination of viral genome equivalents referring to the respective recombinant HFF populations. Viral supernatants were harvested at day 24 p.t. and subjected to IE1-specific qPCR. Calculations were performed in biological sextuplicates per cell population; mean values ± SD are shown. ( B , C ) Statistical analysis was performed using an ordinary two-way ANOVA and post-hoc Tukey correction; ****, p < 0.0001; **, p < 0.01; *, p < 0.1; ns, not significant; no significant differences were noted for values between 1–20 d p.t. ( B ). ( D ) Wb-based expression analysis of the conditionally expressing HFF populations. Recombinant protein expression in the three HFF populations of HFF-UL53, HFF-UL53-Flag and HFF-UL53::sHook1-Flag cells was either uninduced (-) or induced (500 ng/mL Dox). Cells were either infected with HCMV ΔUL53 or remained mock-infected. At 24 h p.t., cells were harvested and lysed. Total lysate samples were subjected to standard Wb analysis using tag-specific or protein-specific monoclonal antibodies as indicated.

Article Snippet: For measurement of HCMV AD169-GFP ΔUL53-positive cells, signals were counted with the ImageXpress ® Pico device (Molecular Devices LLC, San Jose, CA, USA) using CellReporterXpress ® software (version 2.9.3.1183, Molecular Devices LLC) by the system-integrated cell count assay.

Techniques: Virus, Recombinant, Transfection, Fluorescence, Software, Expressing, Infection, Bioprocessing

Journal: Cells

Article Title: ‘Shared-Hook’ and ‘Changed-Hook’ Binding Activities of Herpesviral Core Nuclear Egress Complexes Identified by Random Mutagenesis

doi: 10.3390/cells11244030

Figure Lengend Snippet: Viral replication kinetics of HCMV ΔUL53 and its revertant (HCMV Rev) determined by quantitation of GFP-positive cells and HCMV-specific qPCR on the different recombinant HFF populations. 80,000 inducibly expressing HFFs in 24-well plates were infected with HCMV ΔUL53 or HCMV Rev at a viral dose of 5 × 10 6 genome copies. pUL53, pUL53-Flag or pUL53::sHook1-Flag protein expression was either Dox-induced (+Dox) or remained non-induced (−Dox). ( A ) The number of HCMV-infected cells was measured by detection of GFP signal-positive cells at indicated time points with the CellReporterXpress ® software using the ImageXpress ® Pico device. Values represent 25.04 % of the area of a well and are given as a mean value ± SD of two independently infected wells. ( B ) Viral supernatants were harvested at indicated time points and viral genome equivalents were determined by qPCR. Each value represents the mean ± SD of two independent biological replicates, each measured twice. ( A , B ) Statistical analysis was performed using an ordinary two-way ANOVA and post-hoc Sidak correction; ****, p < 0.0001; ***, p < 0.001; *, p < 0.1; ns, not significant; no significant differences were noted for values between 1–13 d p.i. ( A ) or 1–11 d p.i. ( B ), respectively.

Article Snippet: For measurement of HCMV AD169-GFP ΔUL53-positive cells, signals were counted with the ImageXpress ® Pico device (Molecular Devices LLC, San Jose, CA, USA) using CellReporterXpress ® software (version 2.9.3.1183, Molecular Devices LLC) by the system-integrated cell count assay.

Techniques: Quantitation Assay, Recombinant, Expressing, Infection, Software